Tyrosinemia type I is an autosomal recessive metabolic disease caused by deficiency of the enzyme fumarylacetoacetate hydrolase (FAH). Its main clinical features include progressive liver failure, renal tubular damage, intermittent porphyria like neurologic crisis and the development of hepatocellular carcinomas.The mortality rate is high and death usually occurs in childhood unless liver transplantation is performed. Although this disease is rare in most populations, there are ethnic groups in which the incidence is high, especially in French Canadians. Because of the severity of the disorder and the positive clinical experience with liver transplantation, FAH deficiency is a candidate disorder for gene therapy attempts directed at the liver. Gene transfer into other target tissues such as bone marrow might also be beneficial. However, a natural animal model in which to test such strategies is not available.We propose to create such an animal model by disrupting the gene by homologous recombination in murine embryonic stem cells (ES cells) and subsequently generating mice -homozygous for the FAH gene disruption event. In the initial phase of the project the mouse FAH cDNA will be cloned and employed to isolate and characterize the mouse gene. This information will be used to create a replacement vector capable of disrupting the FAH gene in ES cells by homologous recombination. Chimeric mice will be generated from these ES cells and the mice bread to homozygosity. Hopefully the FAH deficiency mouse will express a phenotype similar to the human disorder and then would be used for 3 main purposes in the future. First it would be utilized to test strategies of delivering a functional FAH cDNA for therapeutic purposes. The main target tissue is liver, but kidney, hemopoietic stem cells and possibly intestine are also potential locations in which expression of the enzyme might alleviate the biochemical defect. Second, the effects of dietary and pharmacologic manipulations could be systematically evaluated for their efficiency. These include phenylalanine, tyrosine and methionine restriction and supplementation with cysteine, glutathione or penicillamine. The third goal is the detailed study of the pathophysiology of tyrosinemia, especially the development of hepatocellular carcinoma. In addition several biochemical features of the disorder such as the elevated methionine and depletion of intracellular glutathione are only poorly understood and could be studied in more detail in the FAH deficient animal.